As a cancer of significant racial disparity, oral cancer has the strongest association with alcohol consumption among all types of cancer. Such an association depends on both the dose and the concentration of alcohol. Our preliminary data have clearly shown that oral administration of alcohol (8%, 15%, 35%, and 50%) promoted oral carcinogenesis in 4-nitroquinoline-1-oxide (4NQ0)-treated mice. Treatment with alcohol of these concentrations mimicked drinking beer, wine, diluted liquor, or straight liquor in humans, respectively. Gene knockout of Nrf2, which regulates expression of multiple anti-oxidative phase 11 enzymes, dramatically enhanced carcinogenesis in the same animal model. Therefore we hypothesize that alcohol promotes oral carcinogenesis by generating oxidative stress in the oral epithelium, and Nrf2-mediated antioxidative response may play a critical role in preventing oral cancer. We plan to test our hypothesis in this grant proposal with the following specific aims: 1. To determine whether alcohol promotes 4NQ0-induced oral carcinogenesis in mice through oxidative stress on the oral epithelium, and whether Cyp2E1 is a major enzyme leading to alcohol-generated oxidative stress. 4NQ0-treated mice will be fed with alcohol (35%), and sacrificed at several time points during the experiment (Week 8, 9, 12, 16, 24). Mouse tongue will be analyzed for histopathology, oxidative damage and Nrf2-mediated antioxidative response. Using wild-type and Cyp2E1-/- mice, we will also determine whether alcohol-generated oxidative stress in the oral epithelium is dependent on Cyp2E1. 2. To examine the role of Nrf2 in alcohol-promoted oxidative stress and oral carcinogenesis using Nrf2-/- and Keap1-/- mice. 4NQ0-treated wild-type and Nrf2-/- mice will be fed with alcohol, and histopathology, oxidative damage, and anti-oxidative response in the oral epithelium will be analyzed to determine the impact of Nrf2-/- on alcohol-promoted oxidative stress and oral carcinogenesis. A short-term experiment on Keap 1-/- mice will determine if genetic activation of Nrf2 may inhibit alcohol-promoted oxidative stress. 3. To investigate whether chemical activators of Nrf2 have chemopreventive effects on alcohol-promoted oral carcinogenesis. 4NQ0-treated wild-type and Nrf2 -/-mice will be fed with alcohol, and treated with two chemically distinct Nrf2 activators from dietary sources, sulforaphane and 3H-1,2-dithiole-3-thione. Histopathology, oxidative damage and antioxidative response in the oral epithelium will be analyzed.